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ABSTRACT The coronavirus SARS-CoV-2 infects host cells by binding to the angiotensin-converting enzyme 2 (ACE2) receptor, which belongs to an anti-inflammatory, anti-thrombotic counter-regulatory arm of the renin-angiotensin system (RAS). ACE2 dysfunction and RAS dysregulation has been explored as a driving force in acute respiratory distress syndrome (ARDS), but data from COVID-19 patients has been inconsistent and inconclusive. We sought to identify disruptions of the classical (ACE)/angiotensin (Ang) II/Ang II type-1 receptor (AT1R) and the counter-regulatory ACE2/Ang 1-7/Mas Receptor (MasR) pathways in patients with COVID-19 and correlate these with severity of infection and markers of inflammation and coagulation. Ang II and Ang 1–7 levels in plasma were measured by enzyme-linked immunosorbent assay (ELISA) for 230 patients, 166 of whom were SARS-CoV-2+. Ang 1–7 was repressed in COVID-19 patients compared to that in SARS-CoV-2 negative outpatient controls. Since the control cohort was less sick than the SARS-CoV-2+ group, this association between decreased Ang 1–7 and COVID-19 cannot be attributed to COVID-19 specifically as opposed to critical illness more generally. Multivariable logistic regression analyses demonstrated that every 10-pg/mL increase in plasma Ang 1–7 was associated with a 3% reduction in the odds of hospitalization (adjusted odds ratio [AOR] 0.97, confidence interval [CI] 0.95 to 0.99) and a 3% reduction in odds of requiring oxygen supplementation (AOR 0.97, CI 0.95 to 0.99) and/or ventilation (AOR 0.97, CI 0.94 to 0.99). Ang 1–7 was also inversely associated with pro-inflammatory cytokines and d-dimer in this patient cohort, suggesting that reduced activity in this protective counter-regulatory arm of the RAS contributes to the hyper-immune response and diffuse coagulation activation documented in COVID-19. IMPORTANCE Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes a unique disease, COVID-19, which ranges in severity from asymptomatic to causing severe respiratory failure and death. Viral transmission throughout the world continues at a high rate despite the development and widespread use of effective vaccines. For those patients who contract COVID-19 and become severely ill, few therapeutic options have been shown to provide benefits and mortality rates are high. Additionally, the pathophysiology underlying COVID-19 disease presentation, progression, and severity is incompletely understood. The significance of our research is in confirming the role of renin-angiotensin system dysfunction in COVID-19 pathogenesis in a large cohort of patients with diverse disease severity and outcomes. Additionally, to our knowledge, this is the first study to pair angiotensin peptide levels with inflammatory and thrombotic markers. These data support the role of ongoing clinical trials examining renin-angiotensin system-targeted therapeutics for the treatment of COVID-19.

Abstract Background Transposable element (TE) sequences are classified into families based on the reconstructed history of replication, and into subfamilies based on more fine-grained features that are often intended to capture family history. We evaluate the reliability of annotation with common subfamilies by assessing the extent to which subfamily annotation is reproducible in replicate copies created by segmental duplications in the human genome, and in homologous copies shared by human and chimpanzee. Results We find that standard methods annotate over 10% of replicates as belonging to different subfamilies, despite the fact that they are expected to be annotated as belonging to the same subfamily. Point mutations and homologous recombination appear to be responsible for some of this discordant annotation (particularly in the young Alu family), but are unlikely to fully explain the annotation unreliability. Conclusions The surprisingly high level of disagreement in subfamily annotation of homologous sequences highlights a need for further research into definition of TE subfamilies, methods for representing subfamily annotation confidence of TE instances, and approaches to better utilizing such nuanced annotation data in downstream analysis.

Evidence of observed negative impacts on natural and human systems from anthropogenic climate change is increasing. However, human systems in particular are dynamic and influenced by multiple drivers and hence identifying an anthropogenic climate signal is challenging. Here we analyze the case of lake Palcacocha in the Andes of Peru, which offers a representative model for other glacier lakes and related risks around the world because it features a dynamic evolution of flood risk driven by physical and socioeconomic factors and processes. Furthermore, it is the object of a prominent climate litigation case, wherein a local Peruvian citizen sued a large German energy producer over risk of flooding from lake Palcacocha. Adopting a conceptual model of cascading impacts and multiple drivers of risk, we first study climatic and other geophysical drivers of flood risk. We find that an anthropogenic signal from flood risk to greenhouse gas emissions is traceable. In parallel, flood risk has been strongly shaped (and increased) by interacting socioeconomic, institutional and cultural processes over the past few decades. The case raises important questions about the differentiation of responsibilities relating to flood risk of both global and local agents, which are, however, difficult to address in cases like Palcacocha, where we reveal a complex network of interlinked global, national and local drivers. Following from this, we outline a normative framework with a differentiated perspective on responsibility, implying that global emitters commit to support strengthening capacities in affected regions and localities and that local institutions and societies engage in local risk reduction measures and policies in collaboration with and driven by local communities.

Abstract Background In recent decades, there has been a growing interest in the impact of insults during pregnancy on postnatal health and disease. It is known that changes in placental development can impact fetal growth and subsequent susceptibility to adult onset diseases; however, a method to collect sufficient placental tissues for both histological and gene expression analyses during gestation without compromising the pregnancy has not been described. The ewe is an established biomedical model for the study of fetal development. Due to its cotyledonary placental type, the sheep has potential for surgical removal of materno-fetal exchange tissues, i.e., placentomes. A novel surgical procedure was developed in well-fed control ewes to excise a single placentome at mid-gestation. Results A follow-up study was performed in a cohort of nutrient-restricted ewes to investigate rapid placental changes in response to undernutrition. The surgery averaged 19 min, and there were no viability differences between control and sham ewes. Nutrient restricted fetuses were smaller than controls (4.7 ± 0.1 kg vs. 5.6 ± 0.2 kg; P

Introduction The target of a class of antiplatelet medicines, P2Y12R inhibitors, exists both on platelets and on brain immune cells (microglia). This protocol aims to describe a causal (based on a counterfactual model) approach for analysing whether P2Y12R inhibitors prescribed for secondary prevention poststroke may increase the risk of cognitive disorder or dementia via their actions on microglia, using real-world evidence.Methods and analysis This will be a cohort study nested within the Swedish National Health and Medical Registers, including all people with incident stroke from 2006 to 2016. We developed directed acyclic graphs to operationalise the causal research question considering potential time-independent and time-dependent confounding, using input from several experts. We developed a study protocol following the components of the target trial approach described by Hernan et al and describe the data structure that would be required in order to make a causal inference. We also describe the statistical approach required to derive the causal estimand associated with this important clinical question; that is, a time-to-event analysis for the development of cognitive disorder or dementia at 1, 2 and 5-year follow-up, based on approaches for competing events to account for the risk of all-cause mortality. Causal effect estimates and the precision in these estimates will be quantified.Ethics and dissemination This study has been approved by the Ethics Committee of the University of Gothenburg and Confidentiality Clearance at Statistics Sweden with Dnr 937-18, and an approved addendum with Dnr 2019-0157. The analysis and interpretation of the results will be heavily reliant on the structure, quality and potential for bias of the databases used. When we implement the protocol, we will consider and document any biases specific to the dataset and conduct appropriate sensitivity analyses. Findings will be disseminated to local stakeholders via conferences, and published in appropriate scientific journals.

Comparison across somatosensory domains, important for clinical and scientific goals, requires prior calibration of impairment severity. Provided test score distributions are comparable across domains, valid comparisons of impairment can be made by reference to score locations in the corresponding distributions (percentile rank or standardized scores). However, this is often not the case. Test score distributions for tactile texture discrimination (n = 174), wrist joint proprioception (n = 112), and haptic object identification (n = 98) obtained from pooled samples of stroke survivors in rehabilitation settings were investigated. The distributions showed substantially different forms, undermining comparative calibration via percentile rank or standardized scores. An alternative approach is to establish comparable locations in the psychophysical score ranges spanning performance from just noticeably impaired to maximally impaired. Several simulation studies and a theoretical analysis were conducted to establish the score distributions expected from completely insensate responders for each domain. Estimates of extreme impairment values suggested by theory, simulation and observed samples were consistent. Using these estimates and previously discovered values for impairment thresholds in each test domain, comparable ranges of impairment from just noticeable to extreme impairment were found. These ranges enable the normalization of the three test scales for comparison in clinical and research settings.

Somatosensory loss post-stroke is common, with touch sensation characteristically impaired. Yet, quantitative, standardized measures of touch discrimination available for clinical use are currently limited. We aimed to characterize touch impairment and re-establish the criterion of abnormality of the Tactile Discrimination Test (TDT) using pooled data and to determine the sensitivity and specificity of briefer test versions. Baseline data from stroke survivors (n = 207) and older neurologically healthy controls (n = 100) assessed on the TDT was extracted. Scores were re-analyzed to determine an updated criterion of impairment and the ability of brief test versions to detect impairment. Updated scoring using an area score was used to calculate the TDT percent maximum area (PMA) score. Touch impairment was common for the contralesional hand (83%) but also present in the ipsilesional hand (42%). The criterion of abnormality was established as 73.1 PMA across older adults and genders. High sensitivity and specificity were found for briefer versions of the TDT (25 vs. 50 trials; 12 or 15 vs. 25 trials), with sensitivity ranging between 91.8 and 96.4% and specificity between 72.5 and 95.0%. Conclusion: Updated criterion of abnormality and the high sensitivity and specificity of brief test versions support the use of the TDT in clinical practice settings.

Introduction: Natural color of dentin is affected by many variables, including anatomical variations, age, how much dentin is exposed, or how much enamel is covering the dentin. Chlorhexidine gluconate (CHX) has been observed to cause tooth staining, especially of exposed dentin. Risk factors for CHX staining include the amount of time for CHX utilization amongst others. Interestingly, the temperature of the rinse when used has been identified as a risk factor. However, no evidence of the effect of temperature is available in the literature. The purpose of this study was to determine the effect of temperature on dentin staining due to CHX exposure.Methods: Two studies were done. The first a pilot study at room temperature to determine the time needed to establish staining solutions, a method to evaluate stain intensity, and establish the time needed to stain dentin samples in vitro. The second study exposed dentin samples on a twice daily basis to a 1 min soak in CHX at different temperatures, followed by a period in an unstimulated saliva mixed with black tea mixture. Temperatures tested were 4, 23, 37 and 50°C. Control samples were exposed to only black tea and saliva (no CHX) and tested at 23°C.Results: The pilot study found that the combination of CHX and black tea causes dentin staining. From this data the sample size needed for the second experiment was calculated, requiring 12 samples per group. Sixty dentin samples were divided amongst 5 groups. The data from this study showed significant darkening of the dentin samples over 18 days. The 4 and 23°C CHX rinses resulted in significant staining compared to the control samples. The 37 and 50°C CHX rinses did not stain significantly more than the control samples.Conclusions: Chlorhexidine has the ability to cause tooth staining in the presence of chromogens such as those in black tea. Significant darkening was observed at lower temperatures (4 and 23°C) over 18 days, therefore dental professionals may wish to advise gently warming the CHX rinse toward 37°C prior to use to reduce the risk of staining.

BackgroundImplementing evidence into clinical practice is a key focus of healthcare improvements to reduce unwarranted variation. Dissemination of evidence-based recommendations and knowledge brokering have emerged as potential strategies to achieve evidence implementation by influencing resource allocation decisions. The aim of this study was to determine the effectiveness of these two research implementation strategies to facilitate evidence-informed healthcare management decisions for the provision of inpatient weekend allied health services.Methods and findingsThis multicentre, single-blinded (data collection and analysis), three-group parallel cluster randomised controlled trial with concealed allocation was conducted in Australian and New Zealand hospitals between February 2018 and January 2020. Clustering and randomisation took place at the organisation level where weekend allied health staffing decisions were made (e.g., network of hospitals or single hospital). Hospital wards were nested within these decision-making structures. Three conditions were compared over a 12-month period: (1) usual practice waitlist control; (2) dissemination of written evidence-based practice recommendations; and (3) access to a webinar-based knowledge broker in addition to the recommendations. The primary outcome was the alignment of weekend allied health provision with practice recommendations at the cluster and ward levels, addressing the adoption, penetration, and fidelity to the recommendations. The secondary outcome was mean hospital length of stay at the ward level. Outcomes were collected at baseline and 12 months later. A total of 45 clusters (n = 833 wards) were randomised to either control (n = 15), recommendation (n = 16), or knowledge broker (n = 14) conditions. Four (9%) did not provide follow-up data, and no adverse events were recorded. No significant effect was found with either implementation strategy for the primary outcome at the cluster level (recommendation versus control β 18.11 [95% CI -8,721.81 to 8,758.02] p = 0.997; knowledge broker versus control β 1.24 [95% CI -6,992.60 to 6,995.07] p = 1.000; recommendation versus knowledge broker β -9.12 [95% CI -3,878.39 to 3,860.16] p = 0.996) or ward level (recommendation versus control β 0.01 [95% CI 0.74 to 0.75] p = 0.983; knowledge broker versus control β -0.12 [95% CI -0.54 to 0.30] p = 0.581; recommendation versus knowledge broker β -0.19 [-1.04 to 0.65] p = 0.651). There was no significant effect between strategies for the secondary outcome at ward level (recommendation versus control β 2.19 [95% CI -1.36 to 5.74] p = 0.219; knowledge broker versus control β -0.55 [95% CI -1.16 to 0.06] p = 0.075; recommendation versus knowledge broker β -3.75 [95% CI -8.33 to 0.82] p = 0.102). None of the control or knowledge broker clusters transitioned to partial or full alignment with the recommendations. Three (20%) of the clusters who only received the written recommendations transitioned from nonalignment to partial alignment. Limitations include underpowering at the cluster level sample due to the grouping of multiple geographically distinct hospitals to avoid contamination.ConclusionsOwing to a lack of power at the cluster level, this trial was unable to identify a difference between the knowledge broker strategy and dissemination of recommendations compared with usual practice for the promotion of evidence-informed resource allocation to inpatient weekend allied health services. Future research is needed to determine the interactions between different implementation strategies and healthcare contexts when translating evidence into healthcare practice.Trial registrationAustralian New Zealand Clinical Trials Registry ACTRN12618000029291.

Abstract Background The lysosphingolipid, sphingosine-1-phosphate, is a well-described and potent pro-angiogenic factor. Receptors, as well as the sphingosine phosphorylating enzyme sphingosine kinase 1, are expressed in the placentomes of sheep and the decidua of rodents; however, a function for this signaling pathway during pregnancy has not been established. The objective of this study was to investigate whether sphingosine-1-phosphate promoted angiogenesis within the placentomes of pregnant ewes. Ewes were given daily jugular injections of FTY720 (2-amino-2[2-(− 4-octylphenyl)ethyl]propate-1,3-diol hydrochloride), an S1P analog. Results FTY720 infusion from days 30 to 60 of pregnancy did not alter maternal organ weights nor total number or mass of placentomes, but did alter placentome histoarchitecture. Interdigitation of caruncular crypts and cotyledonary villi was decreased, as was the relative area of cotyledonary tissue within placentomes. Also, the percentage of area occupied by cotyledonary villi per unit of placentome was increased, while the thickness of the caruncular capsule was decreased in ewes treated with FTY720. Further, FTY720 infusion decreased the number and density of blood vessels within caruncular tissue near the placentome capsule where the crypts emerge from the capsule. Finally, FTY720 infusion decreased asparagine and glutamine in amniotic fluid and methionine in allantoic fluid, and decreased the crown rump length of day 60 fetuses. Conclusions While members of the sphingosine-1-phosphate signaling pathway have been characterized within the uteri and placentae of sheep and mice, the present study uses FTY720 to address the influence of S1P signaling on placental development. We present evidence that modulation of the S1P signaling pathway results in the alteration of caruncular vasculature, placentome architecture, abundance of amino acids in allantoic and amniotic fluids, and fetal growth during pregnancy in sheep. The marked morphological changes in placentome histoarchitecture, including alteration in the vasculature, may be relevant to fetal growth and survival. It is somewhat surprising that fetal length was reduced as early as day 60, because fetal growth in sheep is greatest after day 60. The subtle changes observed in the fetuses of ewes exposed to FTY720 may indicate an adaptive response of the fetuses to cope with altered placental morphology.

Although slow-moving landslides represent a substantial hazard, their detailed mechanisms are still comparatively poorly understood. We have conducted a suite of innovative laboratory experiments using novel equipment to simulate a range of porewater pressure and dynamic stress scenarios on samples collected from a slow-moving landslide complex in New Zealand. We have sought to understand how changes in porewater pressure and ground acceleration during earthquakes influence the movement patterns of slow-moving landslides. Our experiments show that during periods of elevated porewater pressure, displacement rates are influenced by two components: first an absolute stress state component (normal effective stress state) and second a transient stress state component (the rate of change of normal effective stress). During dynamic shear cycles, displacement rates are controlled by the extent to which the forces operating at the shear surface exceed the stress state at the yield acceleration point. The results indicate that during strong earthquake accelerations, strain will increase rapidly with relatively minor increases in the out-of-balance forces. Similar behaviour is seen for the generation of movement through increased porewater pressures. Our results show how the mechanisms of shear zone deformation control the movement patterns of large slow-moving translational landslides, and how they may be mobilised by strong earthquakes and significant rain events.

High and low sodium diets are associated with increased blood pressure and cardiovascular morbidity and mortality. The paradoxical response of elevated BP in low salt diets, aka inverse salt sensitivity (ISS), is an understudied vulnerable 11% of the adult population with yet undiscovered etiology. A linear relationship between the number of single nucleotide polymorphisms (SNPs) in the dopamine D2 receptor (DRD2, rs6276 and 6277), and the sodium myo-inositol cotransporter 2 (SLC5A11, rs11074656), as well as decreased expression of these two genes in urine-derived renal proximal tubule cells (uRPTCs) isolated from clinical study participants suggest involvement of these cells in ISS. Insight into this newly discovered paradoxical response to sodium is found by incubating cells in low sodium (LS) conditions that unveil cell physiologic differences that are then reversed by mir-485-5p miRNA blocker transfection and bypassing the genetic defect by DRD2 re-expression. The renin-angiotensin system (RAS) is an important counter-regulatory mechanism to prevent hyponatremia under LS conditions. Oversensitive RAS under LS conditions could partially explain the increased mortality in ISS. Angiotensin-II (AngII, 10 nmol/L) increased sodium transport in uRPTCs to a greater extent in individuals with ISS than SR. Downstream signaling of AngII is verified by identifying lowered expression of nuclear factor erythroid 2-related factor 2 (NRF2), CCCTC-binding factor (CTCF), and manganese-dependent mitochondrial superoxide dismutase (SOD2) only in ISS-derived uRPTCs and not SR-derived uRPTCs when incubated in LS conditions. We conclude that DRD2 and SLC5A11 variants in ISS may cause an increased low sodium sensitivity to AngII and renal sodium reabsorption which can contribute to inverse salt-sensitive hypertension.

Background: Chronic pain and body perception disturbance are common following stroke. It is possible that an interaction exists between pain and body perception disturbance, and that a change in one may influence the other. We therefore investigated the presence of body perception disturbance in individuals with stroke, aiming to determine if a perceived change in hand size contralateral to the stroke lesion is more common in those with chronic pain than in those without. Methods: Stroke survivors (N = 523) completed an online survey that included: stroke details, pain features, and any difference in perceived hand size post-stroke. Results: Individuals with stroke who experienced chronic pain were almost three times as likely as those without chronic pain to perceive their hand as now being a different size (OR = 2.895; 95%CI 1.844, 4.547). Further, those with chronic pain whose pain included the hand were almost twice as likely to perceive altered hand size than those whose pain did not include the hand (OR = 1.862; 95%CI 1.170, 2.962). This was not influenced by hemisphere of lesion (p = 0.190). Conclusions: The results point to a new characteristic of chronic pain in stroke, raising the possibility of body perception disturbance being a rehabilitation target to improve function and pain-related outcomes for stroke survivors.

MXenes are a relatively new and exciting class of two-dimensional materials, which offer a wide range of compositions and excellent properties including ease of dispersibility and metallic conductivity. These properties render them promising candidates for use as fillers in polymer nanocomposites. Herein, we review how MXene processing methods, their volume fraction, and their incorporation into polymer hosts impact the properties and performance of a wide variety of nanocomposites. Several challenges and barriers that remain are discussed.

Objectives Somatosensory loss is common after stroke with one-in-two individuals affected. Although clinical practice guidelines recommend providing somatosensory rehabilitation, this impairment often remains unassessed and untreated. To address the gap between guideline recommendations and clinical practice, this study sought to understand the factors influencing delivery of evidence-based upper limb sensory rehabilitation after stroke.Design Qualitative study involving focus groups and interviews. Data analysis used an inductive approach (thematic analysis) and deductive analysis using implementation theory (the Theoretical Domains Framework and Normalisation Process Theory).Setting Eight healthcare organisations in metropolitan and regional areas of Victoria and New South Wales, Australia.Participants Eighty-seven rehabilitation therapists (79% occupational therapists and 21% physiotherapists) were purposively sampled and participated in a knowledge translation study with staggered recruitment from 2014 to 2018.Results Three types of factors influenced therapists’ delivery of upper limb somatosensory rehabilitation: individual (‘The uncertain, unskilled therapist’), patient (‘Patient understanding and priorities’) and organisational (‘System pressures and resources’). Deductive analysis using implementation theory identified key determinants of practice change, such as opportunities to consolidate new skills, the anticipated benefits of upskilling as a therapy team and the work anticipated by therapists to incorporate a new somatosensory rehabilitation approach.Conclusions Occupational therapists and physiotherapists hold valuable insights towards practice change in somatosensory rehabilitation from the ‘frontline’. Therapists experience barriers to change including a lack of knowledge and skills, lack of resources and organisational pressures. Facilitators for change were identified, including social support and therapists’ perceived legitimacy in using new somatosensory rehabilitation approaches. Results will inform the design of a tailored implementation strategy to increase the use of evidence-based somatosensory rehabilitation in Australia.Trial registration number Australian New Zealand Clinical Trials Registry (ACTRN12615000933550).

Background The COVID-19 pandemic’s first wave in England during spring 2020 resulted in an approximate 50% increase in all-cause mortality. Previously, risk factors such as age and ethnicity, were identified by studying COVID-related deaths only, but these were under-recorded during this period. Objective To use a large electronic primary care database to estimate the impact of risk factors (RFs) on excess mortality in England during the first wave, compared with the impact on total mortality during 2015–19. Methods Medical history, ethnicity, area-based deprivation and vital status data were extracted for an average of 4.8 million patients aged 30–104 years, for each year between 18-March and 19-May over a 6-year period (2015–2020). We used Poisson regression to model total mortality adjusting for age and sex, with interactions between each RF and period (pandemic vs. 2015–19). Total mortality during the pandemic was partitioned into "usual" and "excess" components, assuming 2015–19 rates represented "usual" mortality. The association of each RF with the 2020 "excess" component was derived as the excess mortality ratio (EMR), and compared with the usual mortality ratio (UMR). Results RFs where excess mortality was greatest and notably higher than usual were age >80, non-white ethnicity (e.g., black vs. white EMR = 2.50, 95%CI 1.97–3.18; compared to UMR = 0.92, 95%CI 0.85–1.00), BMI>40, dementia, learning disability, severe mental illness, place of residence (London, care-home, most deprived). By contrast, EMRs were comparable to UMRs for sex. Although some co-morbidities such as cancer produced EMRs significantly below their UMRs, the EMRs were still >1. In contrast current smoking has an EMR below 1 (EMR = 0.80, 95%CI 0.65–0.98) compared to its UMR = 1.64. Conclusions Studying risk factors for excess mortality during the pandemic highlighted differences from studying cause-specific mortality. Our approach illustrates a novel methodology for evaluating a pandemic’s impact by individual risk factor without requiring cause-specific mortality data.

Background In the UK, large-scale electronic primary care datasets can provide up-to-date, accurate epidemiological information on rarer diseases, where specialist diagnoses from hospital discharges and clinic letters are generally well recorded and electronically searchable. Current estimates of the number of people living with neuromuscular disease (NMD) have largely been based on secondary care data sources and lacked direct denominators. Objective To estimate trends in the recording of neuromuscular disease in UK primary care between 2000–2019. Methods The Clinical Practice Research Datalink (CPRD) database was searched electronically to estimate incidence and prevalence rates (per 100,000) for a range of NMDs in each year. To compare trends over time, rates were age standardised to the most recent CPRD population (2019). Results Approximately 13 million patients were actively registered in each year. By 2019, 28,230 active patients had ever received a NMD diagnosis (223.6), which was higher among males (239.0) than females (208.3). The most common classifications were Guillain-Barre syndrome (40.1), myasthenia gravis (33.7), muscular dystrophy (29.5), Charcot-Marie-Tooth (29.5) and inflammatory myopathies (25.0). Since 2000, overall prevalence grew by 63%, with the largest increases seen at older ages (≥65-years). However, overall incidence remained constant, though myasthenia gravis incidence has risen steadily since 2008, while new cases of muscular dystrophy fell over the same period. Conclusions Lifetime recording of many NMDs on primary care records exceed current estimates of people living with these conditions; these are important data for health service and care planning. Temporal trends suggest this number is steadily increasing, and while this may partially be due to better recording, it cannot be simply explained by new cases, as incidence remained constant. The increase in prevalence among older ages suggests increases in life expectancy among those living with NMDs may have occurred.

[This corrects the article DOI: 10.1371/journal.pone.0244423.].

IntroductionOur hands, with their exquisite sensors, work in concert with our sensing brain to extract sensory attributes of objects as we engage in daily activities. One in two people with stroke experience impaired body sensation, with negative impact on hand use and return to previous valued activities. Valid, quantitative tools are critical to measure somatosensory impairment after stroke. The functional Tactile Object Recognition Test (fTORT) is a quantitative measure of tactile (haptic) object recognition designed to test one’s ability to recognize everyday objects across seven sensory attributes using 14 object sets. However, to date, knowledge of the nature of object recognition errors is limited, and the internal consistency of performance across item scores and dimensionality of the measure have not been established.ObjectivesTo describe the original development and construction of the test, characterize the distribution and nature of performance errors after stroke, and to evaluate the internal consistency of item scores and dimensionality of the fTORT.MethodData from existing cohorts of stroke survivors (n = 115) who were assessed on the fTORT quantitative measure of sensory performance were extracted and pooled. Item and scale analyses were conducted on the raw item data. The distribution and type of errors were characterized.ResultsThe 14 item sets of the fTORT form a well-behaved unidimensional scale and demonstrate excellent internal consistency (Cronbach alpha of 0.93). Deletion of any item failed to improve the Cronbach score. Most items displayed a bimodal score distribution, with function and attribute errors (score 0) or correct response (score 3) being most common. A smaller proportion of one- or two-attribute errors occurred. The total score range differentiated performance over a wide range of object recognition impairment.ConclusionUnidimensional scale and similar factor loadings across all items support simple addition of the 14 item scores on the fTORT. Therapists can use the fTORT to quantify impaired tactile object recognition in people with stroke based on the current set of items. New insights on the nature of haptic object recognition impairment after stroke are revealed.

Currently the longitudinal proteomic profile of post-ischemic stroke recovery is relatively unknown with few well-accepted biomarkers or understanding of the biological systems that underpin recovery. We aimed to characterize plasma derived biological pathways associated with recovery during the first year post event using a discovery proteomics workflow coupled with a topological pathway systems biology approach. Blood samples (n = 180, ethylenediaminetetraacetic acid plasma) were collected from a subgroup of 60 first episode stroke survivors from the Australian START study at 3 timepoints: 3–7 days (T1), 3-months (T2) and 12-months (T3) post-stroke. Samples were analyzed by liquid chromatography mass spectrometry using label-free quantification (data available at ProteomeXchange with identifier PXD015006). Differential expression analysis revealed that 29 proteins between T1 and T2, and 33 proteins between T1 and T3 were significantly different, with 18 proteins commonly differentially expressed across the two time periods. Pathway analysis was conducted using Gene Graph Enrichment Analysis on both the Kyoto Encyclopedia of Genes and Genomes and Reactome databases. Pathway analysis revealed that the significantly differentiated proteins between T1 and T2 were consistently found to belong to the complement pathway. Further correlational analyses utilized to examine the changes in regulatory effects of proteins over time identified significant inhibitory regulation of clusterin on complement component 9. Longitudinal post-stroke blood proteomics profiles suggest that the alternative pathway of complement activation remains in a state of higher activation from 3-7 days to 3 months post-stroke, while simultaneously being regulated by clusterin and vitronectin. These findings also suggest that post-stroke induced sterile inflammation and immunosuppression could inhibit recovery within the 3-month window post-stroke.

Background In spontaneously hypertensive rats (SHR) we observed profound myocardial metabolic changes during early hypertension before development of cardiac dysfunction and left ventricular hypertrophy. In this study, we evaluated whether metformin improved myocardial metabolic abnormalities and simultaneously prevented contractile dysfunction and left ventricular hypertrophy in SHR. Methods and Results SHR and control Wistar–Kyoto rats were treated with metformin from 2 to 5 months of age, when SHR hearts exhibit metabolic abnormalities and develop cardiac dysfunction and left ventricular hypertrophy. We evaluated the effect of metformin on myocardial glucose uptake rates with dynamic 2‐[18F] fluoro‐2‐deoxy‐D‐glucose positron emission tomography. We used cardiac MRI in vivo to assess the effect of metformin on ejection fraction, left ventricular mass, and end‐diastolic wall thickness, and also analyzed metabolites, AMP‐activated protein kinase and mammalian target‐of‐rapamycin activities, and mean arterial blood pressure. Metformin‐treated SHR had lower mean arterial blood pressure but remained hypertensive. Cardiac glucose uptake rates, left ventricular mass/tibia length, wall thickness, and circulating free fatty acid levels decreased to normal, and ejection fraction improved in treated SHR. Hearts of treated SHR exhibited increased AMP‐activated protein kinase phosphorylation and reduced mammalian target‐of‐rapamycin activity. Cardiac metabolite profiling demonstrated that metformin decreased fatty acyl carnitines and markers of oxidative stress in SHR. Conclusions Metformin reduced blood pressure, normalized myocardial glucose uptake, prevented left ventricular hypertrophy, and improved cardiac function in SHR. Metformin may exert its effects by normalizing myocardial AMPK and mammalian target‐of‐rapamycin activities, improving fatty acid oxidation, and reducing oxidative stress. Thus, metformin may be a new treatment to prevent or ameliorate chronic hypertension–induced left ventricular hypertrophy.

Bitter taste receptors (T2Rs) are G protein-coupled receptors (GPCRs) expressed in various cell types including ciliated airway epithelial cells and macrophages. T2Rs in these two innate immune cell types are activated by bitter products, including those secreted by Pseudomonas aeruginosa, leading to Ca2+-dependent activation of endothelial nitric oxide (NO) synthase (eNOS). NO enhances mucociliary clearance and has direct antibacterial effects in ciliated epithelial cells. NO also increases phagocytosis by macrophages. Using biochemistry and live-cell imaging, we explored the role of heat shock protein 90 (HSP90) in regulating T2R-dependent NO pathways in primary sinonasal epithelial cells, primary monocyte-derived macrophages, and a human bronchiolar cell line (H441). Immunofluorescence showed that H441 cells express eNOS and T2Rs and that the bitter agonist denatonium benzoate activates NO production in a Ca2+- and HSP90-dependent manner in cells grown either as submerged cultures or at the air–liquid interface. In primary sinonasal epithelial cells, we determined that HSP90 inhibition reduces T2R-stimulated NO production and ciliary beating, which likely limits pathogen clearance. In primary monocyte-derived macrophages, we found that HSP-90 is integral to T2R-stimulated NO production and phagocytosis of FITC-labeled Escherichia coli and pHrodo-Staphylococcus aureus. Our study demonstrates that HSP90 serves as an innate immune modulator by regulating NO production downstream of T2R signaling by augmenting eNOS activation without impairing upstream Ca2+ signaling. These findings suggest that HSP90 plays an important role in airway antibacterial innate immunity and may be an important target in airway diseases such as chronic rhinosinusitis, asthma, or cystic fibrosis.

Salt sensitivity of blood pressure (BP) refers to an increase in BP following an increase in dietary salt, which is associated with increased incidence of cardiovascular disease and early death. However, decreased sodium intake also increases mortality and morbidity. Inverse salt sensitivity (ISS), defined as a paradoxical increase in BP on a low-salt diet, about 11% of the population, may be the cause of this phenomenon. The epithelial sodium channel (ENaC) is a major regulator of sodium reabsorption in the kidney. In this study, human renal tubular epithelial cells (hRTC) were cultured from the urine of phenotyped salt study participants. αENaC expression was significantly lower in ISS than salt resistant (SR) hRTC, while ENaC-like channel activity was dramatically increased by trypsin treatment in ISS cells analyzed by patch clamp. αENaC expression was also decreased under high-salt treatment and increased by aldosterone treatment in ISS cells. Moreover, the αENaC variant, rs4764586, was more prevalent in ISS. In summary, αENaC may be associated with ISS hypertension on low salt. These findings may contribute to understanding the mechanisms of ISS and low salt effect on morbidity and mortality.

The use of antibiotics to treat dairy calves may result in multidrug-resistant extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli. This study investigated fluoroquinolone and macrolide resistance genes among ESBL-producing E. coli isolated from dairy calves. Fresh fecal samples from 147 dairy calves across three age groups were enriched to select for ESBL-producing E. coli. Plasmid-mediated fluoroquinolone (qnrB), macrolide (mph(A)), and beta-lactam (blaCTX-M groups 1 and 9) resistance genes were identified by PCR and gel electrophoresis in ESBL-producing E. coli. Beta-lactamase variants and antibiotic resistance genes were characterized for eight isolates by whole-genome sequencing. Seventy-one (48.3%) samples were positive for ESBL-producing E. coli, with 159 (70.4%) isolates identified as blaCTX-M variant group 1 and 67 (29.6%) isolates as blaCTX-M variant group 9. Resistance gene mph(A) was more commonly associated with blaCTX-M variant group 1, while resistance gene qnrB was more commonly associated with variant group 9. E. coli growth was quantified on antibiotic media for 30 samples: 10 from each age group. Significantly higher quantities of ceftriaxone-resistant E. coli were present in the youngest calves. Results indicate the dominant blaCTX-M groups present in ESBL-producing E. coli may be associated with additional qnrB or mph(A) resistance genes and ESBL-producing E. coli is found in higher abundance in younger calves.

Simple Summary: L-Glutamate (Glu) and L-glutamine (Gln) are abundant amino acids in feedstuffs and ruminants. Dietary Gln is extensively utilized by ruminal microbes, but dietary Glu undergoes little catabolism by these microbes because they do not take up extracellular Glu due to the lack of the necessary transporters. Microbial proteins and dietary Glu exit the rumen into the abomasum and then the small intestine, where proteins undergo hydrolysis to release amino acids (including Glu and Gln) and small peptides for transport into enterocytes. Most dietary Gln escapes the underdeveloped rumen of preruminants, instead entering the abomasum and the small intestine. Within the enterocytes, Glu and Gln are extensively oxidized to provide ATP and are actively used to synthesize glutathione and other amino acids (alanine, ornithine, citrulline, arginine, proline, and aspartate), whereas Gln and aspartate are essential for purine and pyrimidine syntheses. Under normal feeding conditions, all diet- and rumen-derived Glu and Gln are extracted by the small intestine and, therefore, do not enter the portal circulation. De novo synthesis plays a crucial role in maintaining the homeostasis of Glu and Gln in the whole body but may be insufficient for maximal growth performance, production (e.g., lactation and pregnancy), and optimal health in ruminants. Dietary supplementation with Glu or Gln can safely improve the digestive, endocrine, and reproduction functions of ruminants and thus augment health and production parameters. Although both L-glutamate (Glu) and L-glutamine (Gln) have long been considered nutritionally nonessential in ruminants, these two amino acids have enormous nutritional and physiological importance. Results of recent studies revealed that extracellular Gln is extensively degraded by ruminal microbes, but extracellular Glu undergoes little catabolism by these cells due to the near absence of its uptake. Ruminal bacteria hydrolyze Gln to Glu plus ammonia and, intracellularly, use both amino acids for protein synthesis. Microbial proteins and dietary Glu enter the small intestine in ruminants. Both Glu and Gln are the major metabolic fuels and building blocks of proteins, as well as substrates for the syntheses of glutathione and amino acids (alanine, ornithine, citrulline, arginine, proline, and aspartate) in the intestinal mucosa. In addition, Gln and aspartate are essential for purine and pyrimidine syntheses, whereas arginine and proline are necessary for the production of nitric oxide (a major vasodilator) and collagen (the most abundant protein in the body), respectively. Under normal feeding conditions, all diet- and rumen-derived Glu and Gln are extensively utilized by the small intestine and do not enter the portal circulation. Thus, de novo synthesis (e.g., from branched-chain amino acids and α-ketoglutarate) plays a crucial role in the homeostasis of Glu and Gln in the whole body but may be insufficient for maximal growth performance, production (e.g., lactation and pregnancy), and optimal health (particularly intestinal health) in ruminants. This applies to all types of feeding systems used around the world (e.g., rearing on a milk replacer before weaning, pasture-based production, and total mixed rations). Dietary supplementation with the appropriate doses of Glu or Gln [e.g., 0.5 or 1 g/kg body weight (BW)/day, respectively] can safely improve the digestive, endocrine, and reproduction functions of ruminants to enhance their productivity. Both Glu and Gln are truly functional amino acids in the nutrition of ruminants and hold great promise for improving their health and productivity. [ABSTRACT FROM AUTHOR]

One in two survivors experience impairment in touch sensation after stroke. The nature of this impairment is likely associated with changes associated with the functional somatosensory network of the brain; however few studies have examined this. In particular, the impact of lesioned hemisphere has not been investigated. We examined resting state functional connectivity in 28 stroke survivors, 14 with left hemisphere and 14 with right hemisphere lesion, and 14 healthy controls. Contra-lesional hands showed significantly decreased touch discrimination. Whole brain functional connectivity (FC) data was extracted from four seed regions, i.e. primary (S1) and secondary (S2) somatosensory cortices in both hemispheres. Whole brain FC maps and Laterality Indices (LI) were calculated for subgroups. Inter-hemispheric FC was greater in healthy controls compared to the combined stroke cohort from the left S1 seed and bilateral S2 seeds. The left lesion subgroup showed decreased FC, relative to controls, from left ipsi-lesional S1 to contra-lesional S1 and to distributed temporal, occipital and parietal regions. In comparison, the right lesion group showed decreased connectivity from contra-lesional left S1 and bilateral S2 to ipsi-lesional parietal operculum (S2), and to occipital and temporal regions. The right lesion group also showed increased intra-hemispheric FC from ipsi-lesional right S1 to inferior parietal regions compared to controls. In comparison to the left lesion group, those with right lesion showed greater intra-hemispheric connectivity from left S1 to left parietal and occipital regions and from right S1 to right angular and parietal regions. Laterality Indices were significantly greater for stroke subgroups relative to matched controls for contra-lesional S1 (left lesion group) and contra-lesional S2 (both groups). We provide evidence of altered functional connectivity within the somatosensory network, across both hemispheres, and to other networks in stroke survivors with impaired touch sensation. Hemisphere of lesion was associated with different patterns of altered functional connectivity within the somatosensory network and with related function was associated with different patterns of altered functional connectivity within the somatosensory network and with related functional networks. Keywords: Brain imaging, Resting state fMRI, Somatosensory, Tactile, Stroke, Laterality, Functional connectivity

Abstract Background The treatment of somatosensory loss in the upper limb after stroke has been historically overshadowed by therapy focused on motor recovery. A double-blind randomized controlled trial has demonstrated the effectiveness of SENSe (Study of the Effectiveness of Neurorehabilitation on Sensation) therapy to retrain somatosensory discrimination after stroke. Given the acknowledged prevalence of upper limb sensory loss after stroke and the evidence-practice gap that exists in this area, effort is required to translate the published research to clinical practice. The aim of this study is to determine whether evidence-based knowledge translation strategies change the practice of occupational therapists and physiotherapists in the assessment and treatment of sensory loss of the upper limb after stroke to improve patient outcomes. Method/design A pragmatic, before-after study design involving eight (n = 8) Australian health organizations, specifically sub-acute and community rehabilitation facilities. Stroke survivors (n = 144) and occupational therapists and physiotherapists (~10 per site, ~n = 80) will be involved in the study. Stroke survivors will be provided with SENSe therapy or usual care. Occupational therapists and physiotherapists will be provided with a multi-component approach to knowledge translation including i) tailoring of the implementation intervention to site-specific barriers and enablers, ii) interactive group training workshops, iii) establishing and fostering champion therapists and iv) provision of written educational materials and online resources. Outcome measures for occupational therapists and physiotherapists will be pre- and post-implementation questionnaires and audits of medical records. The primary outcome for stroke survivors will be change in upper limb somatosensory function, measured using a standardized composite measure. Discussion This study will provide evidence and a template for knowledge translation in clinical, organizational and policy contexts in stroke rehabilitation. Trial registration Australian New Zealand Clinical Trials Registry (ANZCTR) retrospective registration ACTRN12615000933550 .

Abstract Background Polyamines stimulate DNA transcription and mRNA translation for protein synthesis in trophectoderm cells, as well as proliferation and migration of cells; therefore, they are essential for development and survival of conceptuses (embryo/fetus and placenta). The ovine conceptus produces polyamines via classical and non-classical pathways. In the classical pathway, arginine (Arg) is transformed into ornithine, which is then decarboxylated by ornithine decarboxylase (ODC1) to produce putrescine which is the substrate for the production of spermidine and spermine. In the non-classical pathway, Arg is converted to agmatine (Agm) by arginine decarboxylase (ADC), and Agm is converted to putrescine by agmatinase (AGMAT). Methods Morpholino antisense oligonucleotides (MAOs) were designed and synthesized to inhibit translational initiation of the mRNAs for ODC1 and ADC, in ovine conceptuses. Results The morphologies of MAO control, MAO-ODC1, and MAO-ADC conceptuses were normal. Double knockdown of ODC1 and ADC (MAO-ODC1:ADC) resulted in two phenotypes of conceptuses; 33% of conceptuses appeared to be morphologically and functionally normal (phenotype a) and 67% of the conceptuses presented an abnormal morphology and functionality (phenotype b). Furthermore, MAO-ODC1:ADC (a) conceptuses had greater tissue concentrations of Agm, putrescine, and spermidine than MAO control conceptuses, while MAO-ODC1:ADC (b) conceptuses only had greater tissue concentrations of Agm . Uterine flushes from ewes with MAO-ODC1:ADC (a) had greater amounts of arginine, aspartate, tyrosine, citrulline, lysine, phenylalanine, isoleucine, leucine, and glutamine, while uterine flushes of ewes with MAO-ODC1:ADC (b) conceptuses had lower amount of putrescine, spermidine, spermine, alanine, aspartate, glutamine, tyrosine, phenylalanine, isoleucine, leucine, and lysine. Conclusions The double-knockdown of translation of ODC1 and ADC mRNAs was most detrimental to conceptus development and their production of interferon tau (IFNT). Agm, polyamines, amino acids, and adequate secretion of IFNT are critical for establishment and maintenance of pregnancy during the peri-implantation period of gestation in sheep.

Accumulating evidence shows that brain functional deficits may be impacted by damage to remote brain regions. Recent advances in neuroimaging suggest that stroke impairment can be better predicted based on disruption to brain networks rather than from lesion locations or volumes only. Our aim was to explore the feasibility of predicting post-stroke somatosensory function from brain functional connectivity through the application of machine learning techniques. Somatosensory impairment was measured using the Tactile Discrimination Test. Functional connectivity was employed to model the global brain function. Behavioral measures and MRI were collected at the same timepoint. Two machine learning models (linear regression and support vector regression) were chosen to predict somatosensory impairment from disrupted networks. Along with two feature pools (i.e., low-order and high-order functional connectivity, or low-order functional connectivity only) engineered, four predictive models were built and evaluated in the present study. Forty-three chronic stroke survivors participated this study. Results showed that the regression model employing both low-order and high-order functional connectivity can predict outcomes based on correlation coefficient of r = 0.54 (p = 0.0002). A machine learning predictive approach, involving high- and low-order modelling, is feasible for the prediction of residual somatosensory function in stroke patients using functional brain networks.

Abstract Mechanical stimulation of airway epithelial cells causes apical release of ATP, which increases ciliary beat frequency (CBF) and speeds up mucociliary clearance. The mechanisms responsible for this ATP release are poorly understood. CALHM1, a transmembrane protein with shared structural features to connexins and pannexins, has been implicated in ATP release from taste buds, but it has not been evaluated for a functional role in the airway. In the present study, Calhm1 knockout, Panx1 knockout, and wild-type mouse nasal septal epithelial cells were grown at an air-liquid interface (ALI) and subjected to light mechanical stimulation from an air puff. Apical ATP release was attenuated in Calhm1 knockout cultures following mechanical stimulation at a pressure of 55 mmHg for 50 milliseconds (p

Abstract The G-protein coupled receptor GPR55 has been postulated to serve as a novel cannabinoid receptor. A previous report indicated that GPR55 knockout mice fail to develop mechanical hyperalgesia, suggesting a pro-nociceptive role for GPR55 in the control of nociceptive responding. However, GPR55 knockout mice remain incompletely characterized in models of pathological pain. Here we provide a comprehensive assessment of responses of GPR55 knockout and wild-type mice to mechanical and thermal (heat, cold) stimulation in multiple, mechanistically distinct models of inflammatory and neuropathic pain. Inflammatory sensitization was produced by intraplantar administration of capsaicin, formalin or complete Freund’s adjuvant. No differences in responding were detected between GPR55 knockout and wild-type mice in any model of inflammatory nociception assessed. Neuropathic pain was induced by partial sciatic nerve ligation (which induces hypersensitivity to mechanical, cold and heat stimulation) or by treatment with the chemotherapeutic agent paclitaxel (which induces hypersensitivity to mechanical and cold stimulation only). No differences were observed between GPR55 knockout and wild type mice in either development or maintenance of neuropathic nociception in either neuropathic pain model. In conclusion, genetic deletion of GPR55 did not alter the development of pathological pain in adult mice in any chronic pain model evaluated.

BackgroundPropensity score techniques can reduce confounding and bias in observational studies. Such analyses are able to measure and balance pre-determined covariates between treated and untreated groups, leading to results that can approximate those generated by randomized prospective studies when such trials are not feasible. The most commonly used propensity score -based analytic technique is propensity score matching (PSM). Although PSM popularity has continued to increase in medical literature, improper methodology or methodological reporting may lead to biased interpretation of treatment effects or limited scientific reproducibility and generalizability. In this study, we aim to characterize and assess the quality of PSM methodology reporting in high-impact otolaryngologic literature.MethodsPubMed and Embase based systematic review of the top 20 journals in otolaryngology, as measured by impact factor from the Journal Citations Reports from 2012 to 2018, for articles using PSM analysis throughout their publication history. Eligible articles were reviewed and assessed for quality and reporting of PSM methodology.ResultsOur search yielded 101 studies, of which 92 were eligible for final analysis and review. The proportion of studies utilizing PSM increased significantly over time (p < 0.001). Nearly all studies (96.7%, n = 89) specified the covariates used to calculate propensity scores. Covariate balance was illustrated in 67.4% (n = 62) of studies, most frequently through p-values. A minority (17.4%, n = 16) of studies were found to be fully reproducible according to previously established criteria.ConclusionsWhile PSM analysis is becoming increasingly prevalent in otolaryngologic literature, the quality of PSM methodology reporting can be improved. We provide potential recommendations for authors regarding optimal reporting for analyses using PSM.

PurposeThe aim of this study was to explore experiences of upper limb somatosensory discrimination retraining in persons with stroke.MethodsA qualitative methodology was used within the context of a randomized control trial of somatosensory retraining: the CoNNECT trial. Participants in the CoNNECT trial completed a treatment program, known as SENSe therapy, to retrain upper limb somatosensory discrimination and recognition skills, and use of these skills in personally valued activities. Eight participants were interviewed on their experience of this therapy. Data were analyzed using Interpretative Phenomenological Analysis (IPA).ResultsFive themes represented participants’ experiences of upper limb somatosensory retraining after stroke: (1) loss of sensation and desire to reclaim normality; (2) harnessing positivity in the therapeutic relationship and specialized therapy; (3) facing cognitive and emotional challenges; (4) distinct awareness of gains and differences in bodily sensations; and (5) improved functioning: control and choice in daily performance. Persons with stroke experienced somatosensory retraining as a valuable treatment that provided them with sensory and functional gains.ConclusionUpper limb somatosensory retraining is a treatment that persons with stroke perceived as challenging and rewarding. People who have experienced stroke believed that somatosensory retraining therapy assisted them to improve their sensation, functional arm use, as well as daily performance and participation in life.